Dottorato in GENETICA E BIOLOGIA MOLECOLARE

Titolo della tesi: Modulation of circRNA biogenesis by m6A modification and characterization of circRNAs in Rhabdomyosarcoma.

Epitranscriptomics have arisen as a crucial regulator of different RNA processes; however, its role in circRNA biogenesis is poorly studied. CircRNAs are a covalently closed transcript formed by fusing a downstream 5' splice donor site to an upstream 3' splice acceptor site in a back-splicing event. Nevertheless, the regulation of back splicing is not yet fully discovered. N6-methyladenosine (m6A) is an RNA modification that plays a role in various RNA metabolism activities, including splicing, stability, and mRNA translation. In this work, we demonstrated that circRNAs expression levels are highly regulated by m6A and that the capacity of a subset of them is dependent on their capacity to bind YTHDC1, an m6A reader involved in mRNA splicing. CircRNAs have been involved in the regulation of diverse oncogenic processes. This thesis describes circVAMP3 and circAFF1, circular RNAs overexpressed in Rhabdomyosarcoma tumours. Specifically, we demonstrated that in RH4 cells, a representative cell line of alveolar Rhabdomyosarcoma, circVAMP3 is differentially deregulated to its linear counterpart by METTL3 and YHDC1, indicating that m6A plays an important role in this circRNA metabolism. Moreover, we show how that circVAMP3 knockdown in RH4 cells impairs cell cycle progression through the alteration of the AKT-related pathways. Also, we proved that circAFF1 inhibits the migration capacity of RH4 cells. Overall, these data suggest that these circular RNAs are novel regulators of alveolar rhabdomyosarcoma progression and could be used as putative therapeutic targets in the future.